John Lee

Since the discovery and implication of N-ethylmaleimide-sensitive factor -attachment protein receptor proteins in membrane fusion almost two decades ago, there have been significant efforts to understand their involvement at the molecular level. In the current study, we report for the first time the molecular interaction between full-length recombinant t-SNAREs and v-SNARE present in opposing liposomes, leading to the assembly of a t-/v-SNARE ring complex. Using high-resolution electron microscopy, the electron density maps and 3D topography of the membrane-directed SNARE ring complex was determined at nanometre resolution. Similar to the t-/v-SNARE ring complex formed when 50 nm v-SNARE liposomes meet a t-SNARE-reconstituted planer membrane, SNARE rings are also formed when 50 nm diameter isolated synaptic vesicles meet a t-SNARE-reconstituted planer lipid membrane. Furthermore, the mathematical prediction of the SNARE ring complex size with reasonable accuracy, and the possible mechanism of membrane-directed t-/v-SNARE ring complex assembly, was determined from the study. Therefore in the present study, using both lipososome-reconstituted recombinant t-/v-SNARE proteins, and native v-SNARE present in isolated SV membrane, the membrane-directed molecular assembly of the neuronal SNARE complex was determined for the first time and its size mathematically predicted. These results provide a new molecular understanding of the universal machinery and mechanism of membrane fusion in cells, having fundamental implications in human health and disease. © 2011 The Authors Journal of Cellular and Molecular Medicine © 2011 Foundation for Cellular and Molecular Medicine/Blackwell Publishing Ltd.

© 2014 Zou et al. Nrf2, a central regulator of the cellular defense against oxidative stress and inflammation, participates in modulating hepatocyte proliferation during liver regeneration. It is not clear, however, whether Nrf2 regulates hepatocyte growth, an important cellular mechanism to regain the lost liver mass after partial hepatectomy. To determine this, various analyses were performed in wild-type and Nrf2-null mice following PH. We found that, at 60 h post-PH, the vast majority of hepatocytes lacking Nrf2 reduced their sizes, activated hepatic progenitor markers, depleted HNF4α protein, and downregulated the expression of a group of genes critical for their functions. Thus, the identity of hepatocytes deficient in Nrf2 was transiently but massively impaired in response to liver mass loss. This event was associated with the coupling of protein depletion of hepatic HNF4α, a master regulator of hepatocyte differentiation, and concomitant inactivation of hepatic Akt1 and p70S6K, critical hepatocyte growth signaling molecules. We conclude that Nrf2 participates in maintaining newly regenerated hepatocytes in a fully differentiated state by ensuring proper regulation of HNF4α, Akt1, and p70S6K during liver regeneration.

Prostate cancer chemoprevention represents a relatively new and promising strategy for reducing the immense public health burden of this devastating cancer of men in the United States and Western societies. Chemoprevention is defined as the administration of agents that modulate one or more steps in the multistage carcinogenesis process culminating in invasive adenocarcinoma of the prostate. In 2000, there were an estimated 170,000 new cases of prostate cancer and 31,000 deaths in the United States. During the past decade, the National Cancer Institute organized the chemoprevention research program and began testing the first generation of promising agents -fenretinide [4-HPR], difluoromethylornithine [DFMO], antiandrogens) in high-risk cohorts and launched the first-large scale US phase 3 primary prevention trial, known as Prostate Cancer Prevention Trial, in 18,000 average-risk men treated for 7 years with finasteride or placebo. In the summer of 1998, the NCI Prostate Cancer Progress Review Group Report to the director of NCI was published in response to the leadership of the prostate cancer advocacy community in conjunction with Congress. To further elucidate and address critical issues identified in this report and to develop a research agenda for the newly created Prostate and Urologic Cancer Research Group in the Division of Cancer Prevention at NCI, the NCI organized the workshop "New Clinical Trial Strategies for Prostate Cancer Chemoprevention." The major objectives were to promote understanding and cooperation among the NCI, US Food and Drug Administration, academia, pharmaceutical industry, and the public regarding new opportunities for clinical prevention trials for prostate cancer. The workshop was divided into three concurrent breakout panels and a fourth joint integrative panel. The workshop addressed multiple key areas identified in the PRG report in the following panels: Molecular Targets and Promising Agents in Clinical Development; Intermediate Endpoint Biomarkers for Prevention Trials; High-Risk Study Populations for Prevention Trials, and Preventive Clinical Trial Designs and Regulatory Issues. Expert panelists were drawn from leading academic, pharmaceutical, and government scientists in basic research and clinical investigation. Key pharmaceutical, biotechnology, academic, and National Institutes of Health scientists presented overviews of their new agents and products in clinical development. Senior FDA physicians from the Center for Drugs and Center for Biologics presented on current standards for new drug and biologic approval for chemoprevention efficacy. Some of the key topics included recent advances in the state of knowledge of promising agents in the clinic based on molecular targets as well as bottlenecks in drug development for pharmaceutical sponsors; strategic modulable biomarkers that can serve as primary endpoints in phase 1/2 trials to assess preventive efficacy; high-risk cohorts with precancer and representative clinical trial designs that are ready for immediate translation into efficient prevention trials, such as Bayesian sequential monitoring for early assessment of biologic activity and factorial designs for assessment of multiagent combinations. Finally, each expert panel generated recommendations for areas of future research emphasizing opportunities and infrastructure needs. © 2001, Elsevier Science Inc.

Drug discovery and development to date has relied on animal models, which are useful but are often expensive, slow, and fail to mimic human physiology. The discovery of human induced pluripotent stem cells has led to the emergence of a new paradigm of drug screening using human and disease-specific organ-like cultures in a dish. Although classical static culture systems are useful for initial screening and toxicity testing, they lack the organization of differentiated iPS cells into microphysiological, organ-like structures deemed necessary for high-content analysis of candidate drugs. One promising approach to produce these organ-like structures is the use of advanced microfluidic systems, which can simulate tissue structure and function at a micron level, and can provide high-throughput testing of different compounds for therapeutic and diagnostic applications. Here, we provide a brief outline on the different approaches, which have been used to engineer in vitro tissue constructs of iPS cell-based myocardium and liver functions on chip. Combining these techniques with iPS cell biology has the potential of reducing the dependence on animal studies for drug toxicity and efficacy screening.

Excited states in Si38,40,42 nuclei have been studied via in-beam γ-ray spectroscopy with multinucleon removal reactions. Intense radioactive beams of S40 and S44 provided at the new facility of the RIKEN Radioactive Isotope Beam Factory enabled γ-γ coincidence measurements. A prominent γ line observed with an energy of 742 keV in Si42 confirms the 2 + state reported in an earlier study. Among the γ lines observed in coincidence with the 2 +→0 + transition, the most probable candidate for the transition from the yrast 4 + state was identified, leading to a 41+ energy of 2173 keV. The energy ratio of 2.93 between the 21+ and 41+ states indicates well-developed deformation in Si42 at N=28 and Z=14. Also for Si38,40 energy ratios with values of 2.09 and 2.56 were obtained. Together with the ratio for Si42, the results show a rapid deformation development of Si isotopes from N=24 to N=28. © 2012 American Physical Society.

We present noninvasive, quantitative in vivo measurements of methemoglobin formation and reduction in a rabbit model using broadband diffuse optical spectroscopy. Broadband DOS combines multifrequency frequency-domain photon migration with time-independent near infrared spectroscopy to quantitatively measure bulk tissue absorption and scattering spectra between 600 nm and 1,000 nm. Tissue concentrations of methemoglobin, deoxyhemoglobin, and oxyhemoglobin were determined from absorption spectra acquired in "real time" during nitrite infusions in nine pathogen-free New Zealand White rabbits. As little as 30 nM [MetHb] changes were detected for levels of [MetHb] that ranged from 0.80 to 5.72 μM, representing 2.2 to 14.9% of the total hemoglobin content. These values agreed well with on-site ex vivo cooximetry data. The reduction of MetHb to functional hemoglobins was also carried out with intravenous injections of methylene blue. As little as 10 nM changes in [MB] were detectable at levels of up to 150 nM in tissue. Our results demonstrate, for the first time, the ability of broadband DOS to noninvasively quantify real-time changes in [MetHb] and four additional chromophore concentrations despite significant overlapping spectral features. These techniques are expected to be useful in evaluating dynamics of drug delivery and therapeutic efficacy in blood chemistry, human, and preclinical animal models. Copyright © 2006 the American Physiological Society.

© Hsiao et al.; licensee BioMed Central.Background: Identifying cellular signaling pathways that become corrupted in the presence of androgens that increase the metastatic potential of organ-confined tumor cells is critical to devising strategies capable of attenuating the metastatic progression of hormone-naïve, organ-confined tumors. In localized prostate cancers, gene fusions that place ETS-family transcription factors under the control of androgens drive gene expression programs that increase the invasiveness of organ-confined tumor cells. C-X-C chemokine receptor type 4 is a downstream target of ERG, whose upregulation in prostate-tumor cells contributes to their migration from the prostate gland. Recent evidence suggests that CXCR4-mediated proliferation and metastasis of tumor cells is regulated by CXCR7 through its scavenging of chemokine CXCL12. However, the role of androgens in regulating CXCR4-mediated motility with respect to CXCR7 function in prostate-cancer cells remains unclear. Methods: Immunocytochemistry, western blot, and affinity-purification analyses were used to study how androgens influenced the expression, subcellular localization, and function of CXCR7, CXCR4, and androgen receptor in LNCaP prostate-tumor cells. Moreover, luciferase assays and quantitative polymerase chain reaction were used to study how chemokines CXCL11 and CXCL12 regulate androgen-regulated genes in LNCaP prostate-tumor cells. Lastly, cell motility assays were carried out to determine how androgens influenced CXCR4-dependent motility through CXCL12. Results: Here we show that, in the LNCaP prostate-tumor cell line, androgens coordinate the expression of CXCR4 and CXCR7, thereby promoting CXCL12/CXCR4-mediated cell motility. RNA interference experiments revealed functional interactions between AR and CXCR7 in these cells. Co-localization and affinity-purification experiments support a physical interaction between AR and CXCR7 in LNCaP cells. Unexpectedly, CXCR7 resided in the nuclear compartment and modulated AR-mediated transcription. Moreover, androgen-mediated cell motility correlated positively with the co-localization of CXCR4 and CXCR7 receptors, suggesting that cell migration may be linked to functional CXCR4/CXCR7 heterodimers. Lastly, CXCL12-mediated cell motility was CXCR7-dependent, with CXCR7 expression required for optimal expression of CXCR4 protein. Conclusions: Overall, our results suggest that inhibition of CXCR7 function might decrease the metastatic potential of organ-confined prostate cancers.

© 2015 American Psychological Association.Objective: To determine if a treatment for interepisode bipolar disorder I patients with insomnia improves mood state, sleep, and functioning. Method: Alongside psychiatric care, interepisode bipolar disorder I participants with insomnia were randomly allocated to a bipolar disorder-specific modification of cognitive behavior therapy for insomnia or psychoeducation as a comparison condition. Outcomes were assessed at baseline, the end of 8 sessions of treatment, and 6 months later. This pilot was conducted to determine initial feasibility and generate effect size estimates. Results: During the 6-month follow-up, the CBTI-BP group had fewer days in a bipolar episode relative to the PE group. The CBTI-BP group also experienced a significantly lower hypomania/mania relapse rate and a marginally lower overall mood episode relapse rate compared with the PE group. Relative to PE, CBTI-BP reduced insomnia severity and led to higher rates of insomnia remission at posttreatment and marginally higher rates at 6 months. Both CBTI-BP and PE showed statistically significant improvement on selected sleep and functional impairment measures. The effects of treatment were well sustained through follow-up for most outcomes, although some decline on secondary sleep benefits was observed. Conclusions: CBTI-BP was associated with reduced risk of mood episode relapse and improved sleep and functioning on certain outcomes in bipolar disorder. Hence, sleep disturbance appears to be an important pathway contributing to bipolar disorder. The need to develop bipolar disorder-specific sleep diary scoring standards is highlighted.

© 2015 Elsevier Inc. β-adrenergic signaling pathways mediate key aspects of cardiac function. Its dysregulation is associated with a range of cardiac diseases, including dilated cardiomyopathy. Previously, we established an iPSC model of familial DCM from patients with a mutation in TNNT2, a sarcomeric protein. Here, we found that the β-adrenergic agonist isoproterenol induced mature β-adrenergic signaling in iPSC-derived cardiomyocytes but that this pathway was blunted in DCM iPSC-CMs. Although expression levels of several β-adrenergic signaling components were unaltered between control and DCM iPSC-CMs, we found that phosphodiesterases 2A and PDE3A were upregulated in DCM iPSC-CMs and that PDE2A was also upregulated in DCM patient tissue. We further discovered increased nuclear localization of mutant TNNT2 and epigenetic modifications of PDE genes in both DCM iPSC-CMs and patient tissue. Notably, pharmacologic inhibition of PDE2A and PDE3A restored cAMP levels and ameliorated the impaired β-adrenergic signaling of DCM iPSC-CMs, suggesting therapeutic potential. In this paper, Wu etal. profiled the β-adrenergic signaling properties in human iPSC-CMs and demonstrated novel epigenetic mechanisms that underlie the compromised β-adrenergic signaling in DCM, a common cause of heart failure and cardiac transplantation. These results enhance our understanding of DCM pathogenesis and may uncover new therapeutic targets.

We argue that there are mutually beneficial connections to be made between ideas in argumentation theory and the philosophy of mathematics, and that these connections can be suggested via the process of producing computational models of theories in these domains. We discuss Lakatos’s work (Proofs and Refutations, 1976) in which he championed the informal nature of mathematics, and our computational representation of his theory. In particular, we outline our representation of Cauchy’s proof of Euler’s conjecture, in which we use work by Haggith on argumentation structures, and identify connections between these structures and Lakatos’s methods.

Theories of diagrams and diagrammatic reasoning typically seek to account for either the formal semantics of diagrams, or for the advantages which diagrammatic representations hold for the reasoner over other forms of representation. Regrettably, almost no theory exists which accounts for both of these issues together, nor how they affect one another. We do not attempt to provide such an account here. We do, however, seek to lay out larger context than is generally used for examining the processes of using diagrams in reasoning or communication. A context in which detailed studies of sub-problems, such as the formal semantics or cognitive impact of specific diagrammatic systems, may be embedded.Accounts of the embedding of sentential logics in the computational processes of reasoners and communicators are relatively well developed from several decades of research in AI. Analogies between the sentential and the graphical cases are quite revealing about both similarities and differences. To provide a structure for the 'grand context' of diagrammatic representation and reasoning, and to clarify the relations between its component problems, we examine carefully these analogies and the decomposition they provide of subproblems for analysing diagrammatic reasoning.